title: Genome-wide map of m6A circRNAs identified in mice model of severe acute pancreatitis

city: Chengdu

first: jun

last: wu

state: Sichuan

address: 270 Tianhuan Road, Rongdu Avenue, Chengdu City, Sichuan Province

department: Laboratory of Basic Medicine

institute: The General Hospital of Western Theater Command

summary: Severe acute pancreatitis (SAP) is the most serious type of pancreatitis with high morbidity and mortality. The underlying pathophysiological mechanism of SAP is complicated and lacking in effective therapeutic options in clinic. In recent years, circular RNAs (circRNAs) are found to be N6-methyladenosine (m6A)-modified and m6A modification of circRNAs plays important roles in physiological and pathological processes. However, the role of m6A modification of circRNAs in SAP remains unknown. Here, we aim to identify differentially expressed m6A circRNAs in SAP and to determine their biological significance and potential mechanisms in SAP. Firstly, we identified 903 m6A peaks that distribute on 781 circRNAs in SAP and control groups. Among them, 57 circRNAs with differentially expressed m6A peaks were identified, of which 32 were upregulated and 25 were downregulated. The total m6A level of circRNAs was reduced compared with control group. Moreover, the function analysis of these m6A circRNAs in SAP found that some important pathways involved in the pathogenesis of SAP, such as protein digestion and regulation of autophagy. In m6A circRNA-microRNA networks, several important miRNAs involved in the occurrence and development of SAP were found to bind to these m6A circRNAs potentially, such as miR-24-3p, miR-26a, miR-92b, miR-216b, miR-324-5p and miR-762. Notably, ALKBH5 was found to be upregulated in SAP. In conclusion, these results demonstrated that m6A modification of circRNAs was widely existed and may play important roles in the pathogenesis of SAP. Our findings provide novel insights regarding understanding the pathophysiological mechanism of SAP and seeking new therapeutic targets for SAP.

overall_design: Exploring the genome-wide map of m6A-modified circRNAs from 3 severe acute pancreatitis mice and 3 control mice

first: Lijun

last: Tang

first: Hongyu

last: Sun

first: Jun

last: Wu

first: Xiaohui

last: Yuan


title: Postnatal day 1.5 small intestine transcriptome analysis in wild type and ASE-/- mice

status: Public on Nov 01 2021

city: Ithaca

first: Natasza

last: Kurpios

state: NY

address: Vet Medical Center, 930 Campus Road

country: USA

institute: Cornell University

type: Expression profiling by high throughput sequencing

summary: By comparing the small intestine transcriptomes of 4 wild type and 3 ASE-/- mice, we identified several oxidative balance related genes are exressed differentially between WT and ASE-/- in postnatal day 1.5 small intestine when dietary nutrient uptake became vital to support neonatal physiology. We here discover that the asymmetry enhancer of Pitx2 is critical for proper redox homeostasis.This is in agreement with the previous discovery that Pitx2 is important for cellular oxidative balance and is therefore important in cellular maturation and differentiation.

overall_design: Intestine mRNA profiles of 1.5-day old wild type (WT) and ASE-/- mice

middle: A


title: CNS antigen-specific control of early B-lineage development in the meninges

city: Hangzhou

first: Heping

last: Xu

state: Zhejiang

address: 18 Shilongshan Road

country: China

institute: Westlake University

summary: In this work we report that early developing B cells present in the meninges of mice at all ages. Single cell RNA-sequencing (scRNA-seq) analysis revealed a consecutive trajectory of meningeal developing B cells in mice and non-human primates (NHPs). Parabiosis together with lineage tracing of hematopoietic stem cells (HSCs) showed that meningeal developing B cells are continuously replenished from the HSC-derived progenitors via a circulation-independent route. Importantly, autoreactive immature B cells which recognize myelin oligodendrocyte glycoprotein (MOG), a central nervous system (CNS)-specific antigen, are eliminated from the meninges but not BM. Furthermore, genetic deletion of MOG tolerated the development/presence/residence of self-reactive B cells in the meninges. Thus, we propose that meninges function as a unique reservoir for B cell development, allowing in situ negative selection of CNS-antigen-autoreactive B cell clones to ensure a local non-self-reactive immune repertoire.

sample_organism:
Macaca mulatta
Mus musculus

platform_organism:
Macaca mulatta
Mus musculus

overall_design: Flow-sorted murine meningeal cells (CD45–CD11b–), which were pre-labeled with hashtag antibodies (Biolegend), were pooled and loaded at 30,000 cells per channel. Flow-sorted macaque immune cell population (CD45–CD11b–CD3–) were loaded at 8,000 cells per channel. The cells were then partitioned into the GemCode instrument, where individual cells were lysed and mixed with beads carrying unique barcodes in individual oil droplets. The products were subjected to reverse transcription, emulsion breaking, cDNA amplification, and sample index attachment. Libraries were pair-end (150+ 150 bp) sequenced on a NovaSeq (Illumina).

first: Yan

last: Wang

first: Dianyu

last: Chen

first: Di

first: Danyang

last: He


title: S-adenosylhomocysteine hydrolase regulates anterior patterning in Dugesia japonica

city: Boston

first: Stephen

middle: M

last: Fuchs

state: MA

address: 4 Blackfan Circle

institute: Institute for protein innovation

summary: We report RNA-seq results from planarians of Dugesia japonica species which have developed resistance to a S-adenosyl homocysteine hydrolase inhibitor AdOx after prolonged exposure.

overall_design: Examination of differentially expressed genes in SAHH inhibitor-resistant planarians relative to wild-type individuals.

first: Kristina

last: Reinmets

first: Johanna

last: Bischof

first: Emily

last: Taketa

first: Michael

last: Levin


title: Expression data of Hep3b cells transfected with si-WIPI2 and si-NC

city: Nanjing

first: Xianwen

state: Jiangsu

address: 321 Zhongshan Road

department: Department of Hepatobiliary Surgery

institute: The Affiliated Drum Tower Hospital of Nanjing University Medical School

summary: WIPI2 regulates the growth of human liver cancer cells. We used microarray to detail the global program of gene expression underlying the cell growth of human liver cancer cell line Hep3b cells.

overall_design: Hep3b cells transfected with si-RNA targeting WIPI2 and negative control was isolated and extracted for hybridization on Affymetrix microarrays. We sought to obtain the change of gene expression after WIPI2 knockdown.


title: mRNA expression changes in miR-19a treated cells

city: Shanghai

first: Sun

last: Hai-Xiang

address: 180 Fenglin Road Shanghai China

institute: Zhongshan Hospital

summary: The goal of the subject is to explore the changed expression mRNA  after miR-19a is  knockdown. We transfected Hepatocelluar carcinoma cell line (Huh7) with control vector or miR-19a inhibitor and got a stable cell line. Then we screened the mRNA expression and found the changed genes.

overall_design: 3 KD vs 3 control

first: Haixiang


title: Mitochondrial respiration contributes to the interferon gamma response in antigen presenting cells

city: East Lansing

first: Andrew

last: Olive

state: MI

address: 567 Wilson Road

department: Microbiology and Molecular Genetics

institute: Michigan State University

summary: Antigen presenting cells (APC) act as an integration point where signaling and metabolic pathways collaboratively determine cellular activation state and shape T cell responses. We performed a series of genome-wide knockout screens in interferon gamma (IFNg)-stimulated macrophages, identifying overlapping sets of regulators that control the expression of the T cell stimulatory or inhibitory proteins, MHCII, CD40, and PD-L1. This multi-screen approach defined novel pathways that differentially control these functionally-distinct proteins, and identified mitochondrial function as a global regulator of the IFNg response. Specifically, we report that complex I of the mitochondrial respiratory chain is necessary for signal transduction via the IFNgreceptor, and consequently for both macrophages and dendritic cells to function as APCs. The importance of mitochondrial function for the IFNg response contrasts with the known glycolytic dependency of pattern-recognition receptor stimulation, implicating metabolic state as a fulcrum of innate immunity.

overall_design: Genome-wide CRISPR knockout screen in murine macrophages for regulators of interferon gamma induced MHCII, CD40 or PD-L1

middle: C

last: Kiritsy

middle: J

first: Christopher

last: Sassetti


title: PERK signaling through C/EBPδ contributes to ER stress-induced expression of immunomodulatory and tumor promoting chemokines by cancer cells.

city: Bethesda

first: Thomas

middle: Joshua

last: Meyer

state: Maryland

address: 41 Center Drive, Building 41, Room B620

department: National Cancer Institute

institute: National Institutes of Health

summary: Expression of the transcription factor CEBPD is induced at early stages of the endoplasmic reticulum (ER) stress response. In order to identify the genes modulated by CEBPD during the ER stress response, we transiently silenced CEBPD expression in MDA-MB-435 melanoma cells and isolated mRNA at 6 h of treatment with Thapsigargin or DMSO control. As control, cells were transfected with two different control siRNA oligonucleotides. Results provide insights into which genes are modulated by CEBPD and/or Thapsigargin in MDA-MB-435 cell

overall_design: MDA-MB-435 cells were transfected with two independent control oligos (siNS and siScr) or CEBPD (siCEBPD1 and siCEBPD2) targeting oligos. 48h post transfection, cells were treated with Vehicle (DMSO) or Tg (100nM) for 6h. The experiment was set up as three biological replicates. RNA was isolated and processed for RNAseq analysis. (NEED TO ADD MORE TECHNICAL DETAILS).

first: Namratha

last: Sheshadri

first: Esta

last: Sterneck


title: Age promotes abortive T follicular helper cell differentiation by increasing Notch signalling.

city: Cambridge

first: Felix

last: Krueger

address: Granta Park

department: Bioinformatics

country: United Kingdom

institute: Altos Labs

summary: Ageing profoundly changes our immune system. It is thought to be a driving factor in the morbidity and mortality associated with infectious disease in the elderly. T follicular helper (Tfh) cells are essential for the generation of long-lasting protective immunity yet we do not know how ageing effect the ability to generate these cells. In this study we examined the effect of age on Tfh-precursor formation in both humans. We found that age drives increased formation of these cells. Further investigation showed that T cells from older donors had altered T cell receptor signalling, increased expression of CXCR5. RNA-Sequencing analysis of cells during differentiation highlighted the Notch-associated transcription factor, RBPJ, as important. Importantly, genetic or chemical modulation of the Notch pathway was able to rescue the age-associated increase in CXCR5 expression and aberrant accumulation of Tfh-precursors.  Despite accumulation of Tfh-precursors, increased Notch activation was not sufficient for further differentiation into full Germinal Center (GC)-Tfh cells. This cell-intrinsic failure to make GC-Tfh cells likely contributes to the defective GC responses in ageing.

overall_design: RNA-Sequencing was performed on CD3/CD28 activated CD4+ T cells from 6 young donors and 6 older donors in the presence/absence of IL-12+TGFB. RNA was extracted on day 3 of activation.

first: Louise

last: Webb

first: Michelle

last: Linterman


title: A genetic screen in macrophages identifies new regulators of IFNg-inducible MHCII that contribute to T cell activation

summary: Cytokine-mediated activation of host immunity is central to the control of pathogens. Interferon-gamma (IFNg) is a key cytokine in protective immunity that induces major histocompatibility complex class II molecules (MHCII) to amplify CD4+ T cell activation and effector function. Despite its central role, the dynamic regulation of IFNg-induced MHCII is not well understood. Using a genome-wide CRISPR-Cas9 screen in murine macrophages we globally identified genes that control MHCII surface expression. Mechanistic studies uncovered two parallel pathways of IFNg-mediated MHCII control that require the multifunctional glycogen synthase kinase 3 beta (GSK3b) or the mediator complex subunit MED16. Both pathways control distinct aspects of the IFNg response and are necessary for IFNg-mediated induction of the MHCII transactivator CIITA, MHCII expression, and CD4+ T cell activation. Our results define previously unappreciated regulation of MHCII expression that is required to control CD4+ T cell responses.

overall_design: RNAseq analysis in murine macrophages of CRISPR mutants that regulate IFN gamma responses. We report RNAseq analysis on control (non-targeted) immortalized macrophages or knockout (sgGSK3b or sgMed16) immortalized macrophages generated using CRISPR in the presence or absence of IFN-gamma stimulation. Total RNA was isolated 24 hours following IFN-gamma stimulation. We generated CRISPR targeted cell lines in immortalized C57bl/6 macrophages that were either non-targeting or targeting Med16 or GSK3b. Knockouts were confirmed by TIDE analysis. We examined the differential expression of genes between genotypes and cytokine activation after 24 hours of IFN-gamma treatment. There are biological triplicates for each genotype and condition resulting in 6 samples per genotype with 3 samples without IFN gamma treatment and 3 samples with IFN gamma Treatment. Total RNA was isolated 24 hours after treatment and processed for RNAseq analysis.

first: Laurisa

last: Ankley


title: Total RNA sequencing analysis to measure the effect of Ehmt2 (G9a, Bat8) deficiencies in the placentas of somite-satge matched embryos

city: Grand Rapids

first: Piroska

middle: E

last: Szabó

address: 333 Bostwick ave NE

department: Epigenetics

institute: Van Andel Institute

summary: The experiment was designed to test the role of EHMT2 in developmental regulation of transcription and imprinted gene regulation. We collected samples that allow us to interpret transcription changes due to different genetically induced deficiencies in the Ehmt2 gene encoding the H3K9 methyltransferase EHMT2. The deficiencies included zygotic homozygosity, maternal mutation, and parental (maternal, paternal, and biparental) haploinsufficiencies, and their respective controls. The time of collection was between 8.5 and 9.5 dpc to allow collecting placentas from somite-stage matched embryos and detect transcriptome changes shortly before the death of homozygous mutant embryos at 10.5 dpc.

overall_design: We performed allele-specific and RNA strain-specific RNA-seq analysis of total RNA samples isolated from individual, 8.5-9.5 dpc placentas of somite stage-matched embryos carrying different deficiencies of Ehmt2. The genotypes and parental genotypes regarding the Ehmt2 mutant allele varied to allow detecting robust or subtle RNA changes due to the specific deficiencies in EHMT2 function. Maternal mutant, maternal haploinsufficient, paternal haploinsufficient and control placentas were collected at the 6-somite stage. Zygotic homozygous and wild type placentas were collected at the 6-somite and 12-somite stage to allow measuring transcriptomic differences in the progression of development. Reciprocal mouse crosses between the 129S1 and JF1/Ms mouse strains were used to generate the placentas to allow distingushing the parental alleles. Four replicate samples (2 males and 2 females) were collected for RNAseq analysis from each condition above. Total RNA was deep sequenced to allow detecting non-coding RNAs.

first: Zeng

last: Tie-Bo

first: Pierce

middle: G

last: Nicholas

first: Szabó

last: Piroska


title: Dephosphorylation of the pre-initiation complex during S-phase is critical for origin firing dynamics

first: Fiona

last: Jenkinson

address: Tennis court road

department: Biochemistry

institute: University of Cambridge

type: Genome binding/occupancy profiling by high throughput sequencing

summary: Genome stability requires complete DNA duplication exactly once before division. In eukaryotes, cyclin-dependent kinase (CDK) plays a dual role in this regulation by inhibiting helicase loading factors and activating origin firing. CDK drives initiation by phosphorylation of two substrates, Sld2 and Sld3, forming the transient and limiting intermediate called the pre-initiation complex (pre-IC). The importance and mechanism of dissociation of the pre-IC from origins is not understood. Here we show in the budding yeast Saccharomyces cerevisiae that CDK phosphorylations of Sld3 and Sld2 are specifically and rapidly turned-over during interphase by the PP2A and PP4 phosphatases. Inhibition of dephosphorylation of Sld3/Sld2 causes dramatic defects in replication initiation genome-wide, retention of the pre-IC at origins and cell death. These studies not only provide a mechanism to ensure that Sld3/Sld2 are dephosphorylated before helicase loading factors but also establish a novel positive role for phosphatases in eukaryotic origin firing.

overall_design: Examination of replication progession and CHIP of Mcm4-3HA and Sld2-6HA by high-throughput sequencing of DNA in two strains (swe1∆ osTir1 cdc55-AID sld3-13myc; swe1∆ osTir1 cdc55-AID pph3-AID sld3-R(2A)-13myc) from G1 to S phase.

first: Miguel

last: Santos

first: Philip

last: Zegerman


title: Adult Leptomeninges are Regionally Patterned and Provide Specialized Support to Underlying Brain Tissue

city: Rensselaer

first: Nathan

last: Boles

address: One Discovery Dr

institute: Neural Stem Cell Institute

summary: The leptomeninges envelop the central nervous system and contribute to brain homeostasis by producing trophic factors and regulating entrance of cells, factors and agents, immune responses and cerebrospinal fluid production. We report that leptomeninges from adult mice are regionally patterned. Leptomeninges dissected from anterior or posterior aspects of the forebrain grown in hetero- or homo-typic culture with anterior and posterior cortical cells demonstrate differences in ability to support survival of neuronal subsets and proliferation of progenitors for astrocytes and oligodendrocytes.  Meningeal support changes with age: co-culture with 18-month old leptomeninges reduced numbers of cortical progenitor cells and neurons but increased astrocyte expansion. Analysis of cytokine secretion and single cell RNA-sequencing revealed differences in anterior versus posterior, young versus old meningeal factors and composition. These data demonstrate that adult leptomeninges are regionally patterned in cell composition and functional properties, and suggest that meningeal deficits may contribute to brain aging and disease.

overall_design: A pool of six male C57B6 mice per timepoint (3 or 18 months old) were used to obtain anterior and posterior meningeal cells.

first: Sally

last: Temple


title: H3K27ac and SMARCB1 binding in control and SMARCB1-knockdown cells

city: seoul

first: Jueng Soo

last: You

address: 120 Neungdong-ro

country: South Korea

institute: Konkuk University

summary: This study delineated the downstream activity of oncogenic SMARCB1 and evaluated the genome-wide change of H3K27ac on chromatin.

overall_design: global change of H3K27Ac in control and SMARCB1 knockdown liver cancer cell line (SK-Hep1)

first: Jueng

middle: S

first: Seong

middle: H

last: Hong

first: Keun

last: Son


title: KLK6-related gene expression profile in FaDu cells

city: Heidelberg

first: Chunxuan

last: Shao

address: Im Neuenheimer Feld 280

country: Germany

institute: German Cancer Research Center

summary: Epithelial-to-mesenchymal transition (EMT) is a dynamic process and critically contributes to treatment resistance, cancer cell dissemination and metastasis. Main objective of this study was to investigate the functional interaction between Kallikrein-related peptidase 6 (KLK6) and aldehyde dehydrogenase 1A (ALDH1A)-related retinoic acid (RA) signaling as recent studies reported their impact on molecular features resembling EMT in several malignancies, including head and neck squamous cell carcinoma (HNSCC). In HNSCC cell lines, we demonstrate that KLK6 is not only a downstream target of ALDH1A-related RA signaling, but also acts as an upstream regulator of ALDH1A3 expression. KLK6 and ALDH1A3 co-expression was confirmed in a mouse xenograft model and two independent HNSCC cohorts (HIPO-HNC and TCGA-HNC). KLK6 and ALDH1A3 co-expressed genes in HNSCC, which are also differentially expression in HNSCC cells with silenced KLK6 expression unraveled several subgroups of HNSCC patients with distinct histopathological and clinical features. In summary, our study highlights a new functional circuit with KLK6 and ALDH1A3 as key nodes and unraveled a gene expression signature, which predicts unfavorable prognosis in human papilloma virus (HPV)-negative HNSCC.

overall_design: Two independent cultures (technical replicates) from three FaDu-shKLK6 clones and FaDu-mock controls, respectively, were used for global expression profiling on the Illumina HumanHT-12 V4.0 array platform. Generation and phenotypic characterization of FaDu-mock and FaDu-shKLK6 clones was described elsewhere (Schrader et al., Mol Cancer. 2015 May 20;14:107).

first: Adriana

last: Jou

first: Xavier

middle: P

last: Hostench

first: Jochen

last: Hess


title: Whole Blood Gene Expression Profiling in Patients Undergoing Laparoscopic Colon Cancer Surgery

city: Roskilde

first: Sara

middle: Kehlet

last: Watt

state: .No State

address: Sygehusvej 6, 3

department: Surgery

country: Denmark

institute: Zealand University Hospital, Roskilde

summary:
We performed whole blood gene expression profiling in 26 patients undergoing laparoscopic surgery for colon cancer stage I-III UICC. Saples were taken the day before surgery (-1), day one postoperatively (+1) and day 10 postoperatively (+10).
The aim of the study was to investigate whether gene expression was altered after laparoscopic colon cancer surgery.

overall_design: The study includes 26 patients with gene expression performed at 3 timepoint (=78 gene whole gele-expression analysis). There are no controlls and no replicates included. Name of sample indicates ID for the patient and day of sample.

middle: K

first: Hans

last: Hasselbalch

first: Vibe

last: Skov

first: Lasse

last: Kjær

first: Mads

last: Thomassen

first: Torben

last: Kruse

first: Mark

last: Burton

first: Ismail

last: Gögenur


title: Tiled Capture-C

city: Oxford

first: A Marieke

last: Oudelaar

address: John Radcliffe Hospital

department: MRC Weatherall Institute of Molecular Medicine

institute: University of Oxford

summary: Capture-C interaction profiles from the viewpoints of the promoters of Chac2, Cpeb4 and Fbxw11 in mouse erythroid and ES cells.

overall_design: Capture-C combines 3C library preparation with oligonucleotide capture for the desired viewpoint restriction fragments. Experiments were performed in biological triplicates of primary erythroid and ES cells.

first: A

first: J

last: Telenius

first: D

middle: R

last: Higgs

first: Jim

last: Hughes


title: Chemical Perturbation of CDK9 and RUVBL2 Reveals Insights on Regulation of the Circadian Phase

city: Dallas

first: Joseph

last: Takahashi

state: TX

address: 5323 Harry Hines Blvd., NA4.118

department: HHMI, Neuroscience

institute: University of Texas Southwestern Medical Center

summary: We report the CRY1 ChIPseq data. Mice were intraperitoneally (I.P.) injected with DRB at CT02 (CT: circadian time). DRB (5,6-dichlorobenzimidazone-1-β-D-ribofuranoside) is a well-known inhibitor of the transcription factor CDK9 which has crucial roles in the RNA polymerase II transcriptional pausing. The mouse liver was collected at 0, 2 and 4hr after treatment. Then, the samples were processed as we previously reported (Koike et al, 2012, Science). This study provides the evidence that DRB regulates the CRY1 binding dynamics.

overall_design: Examination of the DRB effects on the Cry1 biding. DMSO(0hr)_DMSO(2hr)_DMSO(4hr)_DRB(0hr)_DRB(2hr)_DRB(4hr)_and_INPUT

first: Noheon

last: Park

first: Eric

last: Zhang


title: Next Generation Sequencing Facilitates Quantitative Analysis of tetracycline (Tet) inducible Scl/tTA-BCR/ABL (BA) transgenic mouse hematopoietic stem-progenitor cells Transcriptomes

first: Zhiwei

last: Chen

address: Room 807, Research and Education Plaza, 197 Ruijin 2nd Road, Luwan District, Shanghai, PRC

department: Shanghai Institute Of Hematology

institute: Shanghai Jiao Tong Universtiy School of Medicine

summary:
Purpose: The goals of this study are to further clarify the regulation mechanism after BCR/ABL expression was induced for three weeks in Scl/tTA-BCR/ABL trangenic mouse hematopoietic stem-progenitor cells. Moreover, integrated analysis of miRNA-mRNA regulatory network was built by combining the transcriptomic data with miRNA microarray data.
Methods: Hematopoietic stem-progenitor cells mRNA profiles of BA mice after BCR/ABL expression induced for 0 week and 3 weeks were generated by deep sequencing, using Illumina Hiseq 2500, hematopoietic stem-progenitor cells of 3 mice were mixed in each group. The sequence reads that passed quality filters were analyzed at the transcript isoform level with TopHat followed by edgeR. qRT–PCR validation was performed using SYBR Green assays.
Results: Using an optimized data analysis workflow, we investigated the regulation of mRNAs in BM LSKs during murine CML progression, GSEA and GO analysis were taken for the RNA-seq data, revealing that apoptosis pathway was downregulated while cell cycle was upregulated. Then, we comblined RNA-seq data with miRNA microarray data and identified integrated regulatory networks. To validate the analysis results, functional assays were performed with BM LSKs of BA mice and 32D-BCR/ABL cell line.
Conclusions: Our study represents mRNA expression profiles  bone marrow (BM) LSKs (Lin-Sca-1+C-kit+) at 0 and 3 weeks after doxycyline withdrawal of BA mice, the analysis identified that several pathways including cell cycle, apoptosis, myeloid differentiation and DNA damage were influenced by BCR/ABL expression.

overall_design: Hematopoietic stem-progenitor cells (lin-c-kit+sca1+, LSK) mRNA profiles of BA mice after BCR/ABL expression inducing for 0 week and 3 weeks were generated by deep sequencing, using Illumina Hiseq 2500,  LSK cells from three mice were mixed in each group ,

first: Qiuhua

last: Huang


title: Expression of genes associated with poor prognosis from hepatocellular carcinoma are increased in peripheral blood from Hmong with active hepatitis B infection

city: Sacramento

first: Clifford

middle: G.

last: Tepper

state: CA

address: 4645 2nd Avenue, Room 2300A

department: Biochemistry and Molecular Medicine

institute: UC Davis School of Medicine

summary: BACKGORUND & AIMS: Hepatocellular carcinoma (HCC) disproportionally affects Asian Americans due to the high prevalence of chronic hepatitis B virus (HBV) infection and among Asian ethnic groups. The Hmong are diagnosed with HCC at a younger age, with more advanced stage disease, and have the highest cause specific mortality of any Asian ethnic group. Poor survival from HCC has also been associated with differential gene expression in HCC tumors leading us to hypothesize that quantitative traits loci lead to differences in the expression of genes in Hmong which reduce their HCC survival. METHODS: We performed bulk RNA-sequencing of whole blood total RNA from HBV-infected Hmong (n=30) and non-Hmong Asians (Chinese (n=46), and Vietnamese (n=19)). RESULTS: No significant differences between groups were identified in demographic, clinical, or viral factors. A total of 1,491 differentially-expressed transcripts (DETs) were identified across the 3 ethnic groups.  When restricted to those with an HBV viral load >2,000 IU/L, 768 DETs were identified and accurately separated individuals by ethincity. Among 273 DETs identified by functional enrichment and pathway analysis and associated with annotated sets of HCC-related genes, 9 were found in 2 gene sets highly expressed in HCC with poor survival. After adjusting for age and sex, expression levels of 4 of these, including CD164, ECT2, HDAC2, and UBE2T, remained significantly elevated in Hmong compared to Chinese and Vietnamese. CONCLUSIONS: These results suggest expression quantitative trait loci predispose Hmong to a more agrressive form of HCC and may explain the poor survival among this Asian ethnic group.

overall_design: The goal of these studies was to utilize comprehensive transcriptomic profiling in order to determine the differences in gene expression associated with hepatocellular carcinoma (HCC) survival in Laotian/Hmong relative to Chinese and Vietnamese. For this study, we analyzed a total of 97 whole blood RNA samples obtained from a patient cohort consisting of Laotian/Hmong (n=30), Chinese (n=46), and Vietnamese (n=19) patients with chronic hepatitis B. Of these, 42 patients were considered to have active disease (i.e., viral load >2,000 IU/ml). Blood was drawn into PAXgene Blood RNA vacutainer tubes (BD Biosciences) and total RNA isolated using the PAXgene Blood RNA Kit (Qiagen). Indexed, directional RNA-Seq libraries were then prepared from total RNA (1.0 ug) using the TruSeq Stranded Total RNA Library Prep Kit with Ribo-Zero Globin (Illumina, San Diego, CA). All libraries (n=97) were pooled and then multiplex sequenced on an Illumina HiSeq 4000 Sequencing System (100bp, paired-end, 95 samples/lane over 8 lanes).

first: Christopher

middle: L

last: Bowlus


title: Genome-wide maps of IRF3 and YAP in gastric cancer cells.

first: Shi

last: Jiao

address: 320 Yue Yang Road

institute: Shanghai Institutes of Biological Sciences

summary: We performed chromatin immunoprecipitation (ChIP) assays with YAP and IRF3 antibodies in HGC-27 cells. Analysis of the distribution of YAP- or IRF3-binding sites relative to genes annotated in the human genome, revealed that only a minute fraction of peaks mapped close (<1 kb) to transcription start sites (TSSs) whereas most peaks were located farther than 10 kb from the closest TSS. A total of 6,277 peaks were identified by both YAP and IRF3 antibodies.

overall_design: Analysis of genome-wide association between IRF3 and YAP.

first: Jingmin

last: Guan


title: Role of Tcf-1 (Tcf7) in DN3 thymocytes

city: Parkville

first: Alexandra

last: Garnham

state: Victoria

address: 1G Royal Parade

country: Australia

institute: Walter and Eliza Hall Institute of Medical Research

summary: Comparison of wild-type and Tcf-1-deficient DN3 thymocytes

overall_design: Two groups of mouse double negative 3 (DN3) thymocytes are compared. The first group comprizes four samples of wild type (WT) DN3 cells, and the second group consists of two samples of Tcf7 knock-out (KO) DN3 cells. There are six samples in total. All samples are taken from individual mice.

first: Lisa

last: Mielke

first: Cyril

last: Seillet

first: Garbrielle

last: Belz


title: Role of Tcf-1 (Tcf7) in thymic NK cells

summary: Comparison of wild-type and Tcf-1-deficient thymic NK cells

overall_design: Two groups of mouse nature killer (NK) thymic cells are compared. The first group comprizes four samples of wild type (WT) thymic NK cells, and the second group consists of two samples of Tcf7 knock-out (KO) thymic NK cells. There are six samples in total. All samples are taken from individual mice.


title: Gene expression analysis of Tcf7-deficient thymocytes and natural killer cells

summary: This SuperSeries is composed of the SubSeries listed below.

overall_design: Refer to individual Series


title: Nucleosomes inhibit intragenic binding of Rap1p and subsequent cryptic transcription [ChIP-seq]

city: Buffalo

first: Michael

last: Buck

address: 701 Ellicott St

institute: SUNY at Buffalo

summary: Nucleosomes are the primary unit of chromatin structure and inhibit key intracellular interactions which regulate how eukaryotic genetic information is read, including transcription factor (TF) binding to DNA. In theory, the sterics of nucleosome-DNA interactions could inhibit TF binding to all nucleosome-bound DNA. In reality, however, nucleosome inhibition of TF binding is variable both across a genome and even within a single nucleosome. Prior studies have demonstrated that nucleosome occupancy, nucleosome positioning, TF binding site (TFBS) affinity & TFBS location can each influence TF targeting in vivo, however, this work has been largely correlative and no study has dissected collective from individual contributions of these important variables. Therefore, to address this deficiency, we have mapped genome-wide changes in chromatin structure and TF binding following experimental nucleosome depletion. Heuristic modeling of results confirmed that chromatin specification of TF targeting is a multivariate, context-specific, process, with most informative features originating from additive interactions between cis and trans variables. Additionally, we found compelling evidence that nucleosomes serve to inhibit intra-genic TF binding, as evidenced by an increasing number of sub-optimal TFBS occupied following nucleosome depletion. The majority of these conditional loci were located within open reading frames and parallel transcriptome analysis revealed their occurrence was linked to generation of intragenic cryptic transcripts. Together results support the idea that chromatin structure has evolved to coordinate specification of regulatory factor targeting & gene expression. Findings have functional relevance to emerging datasets on genetic and epigenetic variation in human disease states.

overall_design: H4 shutoff (UKY403) and isogenic control (UKY412) strains of S. cerevisiae were obtained from the Grunstein lab. Cells were maintained on agar plates containing YPG media (1% yeast extract, 2% peptone, 2% galactose). For time-course experiments, cells were initially inoculated in 5ml YPG liquid media and cultured at 30C. Logarithmically growing cells were then transferred and diluted into larger 1-1.5L YPG cultures for growth overnight at 30C while shaking in containers with 4x equivalent free-air volume. The following morning, mid-log phase cells (OD600 ~0.6-1.2) were collected by centrifugation, washed 2x within 5 minutes with ~250-500ml YP dextrose (YPD), and re-suspended in equivalent volume of YPD media for 0, 2, and 4 hours of continued growth at 30C. After each time-point samples were taken for MNase-seq, ChIP-seq, and RNA-seq studies. RNA-seq samples were immediately flash-frozen in an ethanol-dry ice bath, whereas samples for ChIP and MNase were first cross-linked at room temperature for 30 min with 1% weight/volume formaldehyde prior to flash freezing.

first: Jonathan

last: Bard

first: Jason

last: Rizzo


title: Nucleosomes inhibit intragenic binding of Rap1p and subsequent cryptic transcription [RNA-seq]


title: Nucleosomes inhibit intragenic binding of Rap1p and subsequent cryptic transcription


title: Role of Tcf-1 (Tcf7) in NK cells

summary: Comparison of wild-type and Tcf-1-deficient spleen NK cells

overall_design: Two groups of mouse nature killer (NK) spleen cells are compared. The first group comprizes two samples of wild type (WT) spleen NK cells, and the second group consists of two samples of Tcf7 knock-out (KO) spleen NK cells. There are four samples in total. All samples are taken from individual mice.

first: Alexandra.

first: Gabrielle


title: Identification of viral and host targets of miR-138 by PAR-CLIP analysis of herpes simplex virus 2 infected 293T cells

city: Hangzhou

first: Dongli

last: Pan

address: 866 Yuhangtang Road

institute: Zhejiang University

summary: Like herpes simplex virus 1 (HSV-1) ICP0 mRNA, HSV-2 ICP0 mRNA is predicted to be targeted by a host neuron-specific microRNA, miR-138. This study was designed to confirm the interaction between HSV-2 ICP0 mRNA and miR-138, and to identify other potential viral and host targets of miR-138 during HSV-2 infection. We performed PAR-CLIP on HSV-2 infected 293T cells overexpressing miR-138 in comparisin to control 293T cells. The results confirmed ICP0 as miR-138's targets, and also identified some other potential viral and host targets of miR-138.

overall_design: Using previously generated 293T cells transduced with lentivirus expressing miR-138 and control 293T cells transduced with an empty vector, we infected these cells with HSV-2 for 4 hours at a multiplification of infection (MOI) of 1, and then harvested the cells for PAR-CLIP analysis. Sites with differential reads between the two kinds of cells are identified as potential target sites of miR-138.

first: Donald

last: Coen


title: Gene expression changes induced in mouse bone marrow by placenta growth factor (PlGF) overexpression.

city: Genoa

first: Paolo

last: Uva

address: Via Gerolamo Gaslini, 5

country: Italy

institute: IRCCS Istituto Giannina Gaslini

summary:
Placenta growth factor (PlGF) is a member of the vascular endothelial growth factor family and plays an important role in adult pathological angiogenesis. To further investigate PlGF functions in tumor growth   and metastasis formation, we used transgenic mice overexpressing PlGF.
Recent studies demonstrate that tumor vascularization does not exclusively rely on preexisting vessels but also depends on bone marrow derived progenitor cells. The aim of the present work is to investigate the effect of the PlGF overexpression in the skin on the transcriptional landscape of bone marrow derived cells.

overall_design: We compared the expression profiles of 4 transgenic mice overexpressing the placenta growth factor (PlGF)  with 4 control mice.

first: Antonella

last: Travaglione

first: Cristina

last: Failla


title: An Antagonist of Toll-like Receptor 7 and 9 lowers cholesterol and reduces atherosclerosis in hyperlipidemic mice

summary: The impact of a dual TLR7 and TLR9 antagonist on hypercholesteremia and atherosclerosis was evaluated in hyperlipidemic mice.  Treatment with antagonist induced a dose dependent reduction of total cholesterol (TC) and LDL-cholesterol.  Changes in Adiponectin, Leptin and free fatty acids levels were observed.  Plaque formation was inhibited in ApoE-/- and LDL-R-/- mice fed high fat diet and treated with the antagonist.  Hepatic and renal steatosis was reduced by the treatment.  Induction of IL-10 expression was inversely correlated with TC levels.  The antagonist induced decrease in hepatic IKKα protein level and enhancement of Akt and Gsk3β phosphorylation.  Treatment of C57BL/6 mice caused increase in hepatic LXR, PPARγ and ABCG1 expression and in stool content of TC.  Antagonist treatment resulted in altered expression of several genes and pathways related to immune system, cholesterol, fatty acid and glucose metabolism.  Data obtained suggest involvement of TLR7 and TLR9 in diet induced hyperlipidemia and atherosclerosis and demonstrate therapeutic potential of TLR7/9 antagonist.

overall_design: Total RNA isolated from C57BL/6 HFD and ApoE-/- HFD mice treated with antagonist compared to untreated mice

first: Fu-Gang

last: Zhu

first: Weiwen

last: Jiang

first: Wieslawa

last: Mentzen

last: Reardon

first: Dong

last: Yu

first: Ekambar

last: Kandimalla

first: Nicola

last: La Monica

first: Sudhir

last: Agrawal


title: Single-cell RNA sequencing reveals a strong connection between Gadd45g upregulation and oncolytic HSV infection in tumor tissue

status: Public on Oct 31 2021

city: Houston

first: Guangsheng

last: Pei

state: Texas

address: 7000 Fanin

institute: UTHealth

summary: The oncolytic effect of virotherapy derives from the intrinsic capability of the applied virus in selectively infecting and killing tumor cells. Although oncolytic viruses of various constructions have been shown to efficiently infect and kill tumor cells in vitro, the efficiency of these viruses to exert the same effect on tumor cells within tumor tissues in vivo has not been extensively investigated. Here we report our studies using single-cell RNA sequencing to comprehensively analyze the gene expression profile of tumor tissues following herpes simplex virus 2-based oncolytic virotherapy. Our data revealed the extent and cell types within the tumor microenvironment that could be infected by the virus. Moreover, we observed changes in the expression of cellular genes, including antiviral genes, in response to viral infection. One notable gene found to be upregulated significantly in oncolytic virus-infected tumor cells was Gadd45g, which is desirable for optimal virus replication. These results not only help reveal the precise infection status of the oncolytic virus in vivo, but also provide insight that may lead to the development of new strategies to further enhance the therapeutic efficacy of oncolytic virotherapy.

overall_design: FusON-H3, HSV-2 based oncolytic virus at 5X10^6 pfu was injected intra-tumorally into 8-10 mm subcutaneous H7, H7-Her2, MC38gp100 and MC38gp100-Her2 tumors in C57BL/6 mice (n=3). The tumors were harvested at 48 hours post-virotherapy, pooled, digested dissaggregated to single cell suspensions. The single cell preparations were then sorted into CD45+ and CD45- cells and mixed at the ratio of 3:1 and processed for scRNA-seq.

first: Divya

last: Ravirala

first: Zhongming

last: Zhao

first: Shaun

last: Zhang


title: A long non-coding RNA in the let-7complex acting as a potent and specific death effector of cancer cells [time course]

city: Basel

first: Tommy

middle: Beat

last: Schlumpf

state: Basel-Stadt

address: Mattenstrasse 26

department: D-BSSE

country: Switzerland

institute: ETH Zürich

summary: The let-7 complex in Drosophila encodes three evolutionarily conserved microRNAs: miR-100, let-7, and miR-125. These act as heterochronic genes in regulating developmental timing in response to the steroid hormone ecdysone and play important roles in cell differentiation. Here we identify two additional long non-coding RNAs in the let-7 complex, we named let-A and let-B. Both are transcribed in the large first intron of the primary RNA encoding the microRNAs. We show these RNAs to be sequentially expressed in early pupal stages in response to ecdysone signaling, albeit exhibiting a different expression pattern compared to the microRNA let-7. Surprisingly, ectopic expression of let-A in Drosophila cancer cells induces rapid cell death. Dead cells further release RNA molecules in the medium that is becoming toxic to other cancer cells. In vivo grown tumors lose their tumorigenicity after being incubated in the let-A induced medium. Moreover, feeding flies carrying transplanted tumor cells with such induced medium leads to reduced growth of tumors in a subset of hosts. Our results uncover a new lncRNA which can act as a potent and specific cell death effector for Drosophila tumor cells.

overall_design: Determination of gene expression profile of ph505 cells before and at 8, 16 and 24 hours after μM 20-hydroxy-ecdysone exposure with 4 biological samples each.

first: Tosca

last: Birbaumer

middle: B

first: Makiko

last: Seimiya

first: Yanrui

first: Renato

last: Paro


title: AGING REGULATES VIRUS-TRIGGERED ASTHMATIC AIRWAY PATHOLOGY

city: St Louis

first: Jeffery

last: Haspel

state: MO

address: 660 S Euclid Avenue

institute: Washington University, St Louis

summary: To understand why asthma remit with aging, we exposed mice across a range of ages to viral and allergic triggers of asthma exacerbations and inflammatory airway pathology.  We found that pathology induced by Sendai virus (SeV) or influenza A virus (IAV) occurred selectively in juvenile mice in a microbiome-independent manner, while the same phenotypes induced by allergens were relatively insensitive to age.  Age-specific responses to SeV included a juvenile bias towards type-2 airway inflammation that emerged early in infection and was lost with maturation.  With aging, we observed progressive transcriptional changes to alveolar macrophages (AMs) including the acquisition of high-level MHC-II expression.  Importantly, depleting AMs before SeV infection canceled the protective effects of maturity on post-viral airway pathology.   Thus, age-related changes to the lung immune micro-environment alter host responses to viruses and may drive childhood asthma remission.

overall_design: We induced asthmatic airway phenotypes in mice across a span of ages using viral and allergic triggers of inflammatory airway pathology. We compared data of airway mechanics, airway inflammation, whole lung mass cytometry, single cell RNA seq and bulk RNA extracted from mice bronchoalveolar lavage.

first: Jeffrey

first: Guy

last: Hazan

first: Carrie

last: Gierasch

first: Anna

last: Ehlers


title: A long non-coding RNA in the let-7complex acting as a potent and specific death effector of cancer cells


title: Real-time quantitative PCR analysis of EV-associated miRNAs from Human Tconv cell

city: Napoli

first: Paola

last: De Candia

address: via Pansini 5

institute: UNIVERSITA' DEGLI STUDI DI NAPOLI FEDERICO II - Dipartimento di Medicina molecolare e Biotecnologie mediche

summary: MicroRNAs (miRNAs) are small non-coding molecules targeting messenger RNAs and inhibiting protein translation. Regulated by dynamic micro-environmental cues, miRNAs in turn modulate key biological processes, including cell growth and development, energy utilization and homeostasis. In particular, miRNAs control the differentiation, survival and activation of CD4+ T conventional (Tconv) cells, key players of the adaptive immunity and miRNA-mediated gene expression regulation contributes to the physiological response to infections and the pathological loss of immune homeostasis in autoimmunity. Upon T cell receptor (TCR) stimulation, the described global miRNA quantitative decrease occurring in T cells is believed to promote the acquisition of effector functions by relaxing the post-transcriptional repression of genes associated to proliferation and cell activity. While miRNAs were initially thought to get down-regulated uniquely by intracellular degradation, miRNA secretion via extracellular vesicles (EVs) represents an additional mechanism of rapid down-regulation. By focusing on molecular interactions by means of graph theory, we have found that miRNAs released by TCR stimulated Tconv cells are significantly enriched for targeting transcripts up-regulated upon stimulation, including those encoding for crucial proteins associated to Tconv cell activation and function. Based on this computational approach, we present our perspective based on the following hypothesis: a stimulated Tconv cell will release miRNAs targeting genes associated to the effector function in the extracellular space in association with EVs, which will thus possess a suppressive potential toward other Tconv cells in the paracrine environment. We also propose possible future directions of investigation aimed at taking advantage of these phenomena to control Tconv cell effector function in health and autoimmunity.

overall_design:
Human Tconv cell-derived EVs were isolated from conditioned media using size exclusion chromatography (Exo-spin™ columns, Cell Guidance) according to manufacturer’s protocol. Isolated EV eluate was total RNA extracted using RNAqueous-4PCR Kit (Ambion, U.S.A.) and a fixed volume of eluted RNA sample was used as input for reverse transcription reaction by miRCURY-LNA RT Kit according to manufacturer’s instruction (Qiagen, U.S.A.). EV-associated miRNAs (n = 752) were profiled by using the complete human miRCURY LNA miRNA panel I+II (V5, Qiagen, U.S.A.).
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Authors state Since we analyzed only test condition, and since the study purpose was the identification of miRNA expressed, normalized data matrix and fold change values are not reported.

first: Silvia

last: Garavelli

first: Dario

last: Di Silvestre


title: Cellular basis of omentum activation and expansion revealed by single-cell RNA sequencing using a parabiosis model

city: Bunkyo-ku

first: Kyohei

last: Fujita

state: Tokyo

address: 1-5-45, Yushima

country: Japan

institute: Institute of Science Tokyo

summary: Although the physiological function of the omentum remains elusive, it has been proposed that it plays an important role in fat storage, immune regulation, and regeneration of injured tissues and organs. Although the omentum undergoes expansion upon activation, reports on the accurate assessment of increased cell types and the origin of the increased cells remain limited. To investigate this aspect, the omenta of parabiotic mice were subjected to activation using distinct fluorescent markers and single-cell RNA sequencing (scRNA-seq) was performed to identify circulation-derived omental cells. We found that a considerable number of circulating cells contributed to the activation of the omentum. The omental cells derived from circulating cells exhibited morphological features similar to those of fibroblasts. scRNA-seq revealed the existence of a novel cell population that co-expressed macrophage and fibroblast markers in the activated omentum, suggesting that it corresponded to circulating macrophage-derived fibroblast-like cells. Lineage tracing experiments revealed that most fibroblasts in the activated omentum were not derived from WT1-positive mesenchymal progenitors. The cell cluster also expressed various chemokine genes, indicating its role in the activation and recruitment of immune cells. These results provide important information regarding the activation of the omentum.

overall_design: mRNA profiles of omental cells from parabiotic Rosa26-ECFP/+ mice paired with Rosa26-EGFP/+ mice

first: Kazuhiro

last: Ishigaki

first: Keiki

last: Kumano

first: Hiroo

last: Ueno


title: transcript analysis of strains with or without plasmid containing 40 genes in three different culture conditions

city: Shenzhen

first: Li

last: Cheng

address: Xueyuan Boulevard

department: Institute of Synthetic Biology

institute: Shenzhen Institutes of Advanced Technology, Chinese Academy of Sciences

summary:
Purpose: To investigate the influence of the 40 genes on plasmid, we compared the transcript varies between the engineered strains and the wild-type strain in synthetic medium with different carbon sources (i.e., glucose, galactose and ethanol/glycerol media).
Methods:  Total mRNA profiles of the engineered strains and the wild-type strain were generated by deep sequencing, in triplicate, using BGIseq500. The sequence reads that passed quality filters were analyzed.
Results:  RNA sequencing results revealed that the transcript patterns were influenced dramatically by the carbon sources and there was no significant difference between the wild-type and engineered strains (that is, any change amounted to less than 10%)

overall_design: Total mRNA profiles of the engineered strains and the wild-type strain which grew to log phase and collected.

first: Junbiao

last: Dai


title: Nucleosomes inhibit intragenic binding of Rap1p and subsequent cryptic transcription [MNase-seq]

first: Michael

last: Buck

first: Jonathan

last: Bard

first: Jason

last: Rizzo


title: Transcriptomics of the HPG axis from in vitro or in vivo-produced dairy male calves

city: Utrecht

first: Maria

middle: Belen

last: Rabaglino

address: Yalelaan 7

department: Population Health Science

country: Netherlands

institute: Utrecht University

summary: In cattle, two of the most common technologies for embryo production are “in vivo” (MOET) or “in vitro” (IVP). IVP can sometimes impact fetal growth, resulting in large offspring, although most IVP calves are born apparently healthy. However, gene regulation changes in the organs involved in the developmental process in phenotypically normal postnatal calves are unknown and could indicate physiological programming. The objective of this study was to compare the transcriptomic modifications in organs of the hypothalamus-pituitary-gonadal and adrenal (HPGA) axes between IVP and MOET male calves of three months of age. RNA extracted from the HPGA organs samples (n=4 per group) were sequenced through RNAseq. The raw files were subjected to a bioinformatic pipeline to determine differentially expressed genes (FDR<0.05) in IVP versus MOET samples and the KEGG pathways enriched by these genes (FDR < 0.1). Pathways related to the reproductive process (GnRH secretion in hypothalamus, GnRH signaling in pituitary and steroidogenesis in testicle) were stimulated in IVP calves, suggesting an early activation of the HPG axis in male IVP calves when compared to MOET counterparts.

overall_design: Samples were obtained from the hypothalamus, pituitary, testicle and adrenal from three-month-old IVP calves (n=4) and control MOET calves (n=4).

first: Jan

last: Bojsen-Møller Secher

first: Marc

middle: A

last: Sirad

first: Poul

last: Hyttel

first: Haja

middle: N

last: Kadarmideen


title: RNA-Sequece Analysis of si-THAP7-AS1 and SI-NC IN 468 CELLS

city: Jinan

first: Haiting

last: Liu

address: West Wenhua Road 44

institute: Shandong University

summary: RNA-seq was used to analyze the differential expression of mRNAs between si-THAP7-AS1 and si-NC468 cells.

overall_design: THAP7-AS1 was knocked down by transfection with THAP7-AS1 Smart Silencer in 468 cells. After 48h, we obtained the samples.

first: Hai-ting

first: Sen

first: Xiang-yu

last: Guo

first: Guo-hao

last: Zhang


title: Transcriptomic analysis of four mouse strains

city: Taipei

first: Dhivya

last: Appan

address: Rm: 1547, 15th Floor, No.1, Sec.1, Ren-ai Rd.

department: Graduate Institute of Brain and Mind Sciences

country: Taiwan

institute: National Taiwan University

summary: The goal of this study is to compare the mouse strains C57BL/6 and CAST/EiJ and their hybrids of forward and reverse mating (F1i and F1r respectively) in behavioral, molecular and electrophysiological level. We examine the effects of parental genotypes on the offspring phenotypes using behavioral studies, electrophysiology and RNA-seq. We show that despite having the same genome, the F1i and the F1r hybrids display different phenotypes, and this may be due to having opposite parental sources of the same genome.

overall_design: Whole brain mRNA profiles of B6, CAST, B6 x CAST (F1i) and CAST x B6 (F1r) mice

first: Hsien-Sung

last: Huang


title: H3K36me3 Reader NPAC/GLYR1 Recruits H3K4 Demethylase LSD2/KDM1B to Modulate Active Gene Transcription

city: Boston

first: Rui

last: Fang

address: 221 Longwood Ave. EBRC 222

institute: Brigham and Women's Hospital

type: Genome binding/occupancy profiling by array

summary: NPAC ChIP were performed by anti-Flag and anti-HA tandem affinity purification from HeLa stably expressing Flag-HA tagged NPAC from pOZ-N vector, and enrichement on chromosome 3, 21, and 22 were determined by chip microarray analysis using Affymatrix HumanTiling 2.0 arrays

overall_design: NPAC ChIP-on-chip were performed in parallel with LSD2 (GSM565104) and enrichment were determined by comparing to input hybridization. The Input and LDS2 ChIP samples from GSM565104 were included here since all were used in the analysis.

first: Yujiang

last: Shi


title: cDNA Microarray 2: Crown developing xylem vs. Base developing xylem

city: Málaga

first: Francisco

middle: R.

last: Cantón

state: Málaga

address: Campus de Teatinos, s/n

department: Biología Molecular y Bioquímica

country: Spain

institute: Universidad de Málaga

summary: Transcriptional analysis comparing the transcriptomes of crown and base developing xylems from adult pine trees. Molecular factors responsible of wood properties can be identified with this strategy.

overall_design: Two-condition experiment Crown Differentiating Xylem vs. Base Differentiating Xylem. Biological replicates: 3 crown xylem, 3 base xylew, harvested from three different individual pine trees. Two technical replicates per array (A and Z). Two pairs of crown and base xylem RNA samples were labeled in the same way (crown xylem with Cy3 and base xylem with Cy5), while for labelling of the third pair dye-swap was performed (crown xylem labelled with Cy5 and base xylem with Cy3)

first: S

middle: M

last: Díaz-Moreno

first: D

last: Pacheco-Villalobos

first: N

last: Fernández-Pozo

first: M

last: Claros

first: F

last: Cánovas


title: RNA Sequencing Analysis of cultured macrophages harvested from the infarcted heart of WT mice and il21 receptor deficient mice (il21r-/-)

city: Chiba City

first: Akihiko

last: Kubota

state: Chiba

address: 1-8-1 Inohana

department: Department of Cardiovascular Medicine

institute: Graduate School of Medicine, Chiba University

summary: Ly6Clow macrophages promote scar formation and prevent early infarct expansion after myocardial infarction (MI). Although CD4+ T cells influence the regulation of Ly6Clow macrophages after MI, the mechanism remains largely unknown. Here, we focused on IL-21 and uncovered its physiological relevance in post-MI hearts. CD4+ T cells harvested from the infarcted heart produce IL-21 upon stimulation, and IL-21 receptor was expressed on Ly6Clo macrophages in the infarcted heart. The survival rate after MI was significantly improved in IL-21-deficient mice compared with WT mice. Moreover, transcriptome analysis of infarcted heart tissue demonstrated that inflammation was persistent in WT mice compared with IL-21-deficient mice. The number of neutrophils was significantly decreased, whereas the number of Ly6Clow macrophages was significantly increased in IL-21-deficient mice. Consistently, IL-21 enhanced the apoptosis of Ly6Clow macrophages. Furthermore, RNA-seq analysis of Ly6Chi and Ly6Clo macrophages stimulated with or without IL-21 for 24 hours revealed that IL-21 induces inflammatory responses in both Ly6Chi and Ly6Clo macrophages. Finally, the treatment with IL-21 receptor Fc protein significantly increased the survival after MI. Thus, the deletion of IL-21 improves survival after MI by preventing Ly6Clo macrophage apoptosis.

overall_design: We collected Ly6Chi macrophages and Ly6Clo macrophages from the infarcted hearts of WT mice and il21 receptor deficient mice (il21r-/-) at 3 days after MI. mRNA profiles from cultured macrophages stimulated with or without IL-21 for 24 hours.

first: Arifumi

last: Iwata

first: Akira

last: Suto

first: Hiroshi

last: Nakajima


title: Expression data from Luteolin-treated human Neural Stem Cells (hNSCs)

city: Tsukuba

first: Mariem

last: Achour

state: Ibaraki

address: 1-1-1 Tennodai

institute: Alliance for Research on the Mediterranean and North Africa

summary:
Gene expression profiling reveals a potential role of Luteolin in  human neuronal stem cells (hNSCs) differentiation .
hNSCs purchased from Gibco were treated with 1 μM verbenalin for 24 hours. Microarray gene expression profiling was conducted for biological replicates of hNSCs cultured in differentiation cell culture medium supplemented with Luteolin for 24 hours  and untreated control cells cultured in differentiation cell cultured medium .

overall_design: RNA was extracted using Isogen (Nippon Gene Co. Ltd., Toyama, Japan). The integrity of RNA was quantified using NanoDrop 2000 spectrophotometer (Thermo Scientific, Wilmington, DE, USA). RNA samples were prepared for gene expression profiling analysis with GeneChip® 3' Expression Arrays using 3' IVT PLUS Reagent Kit (Affymetrix Inc., Santa Clara, CA, USA). One hundred  ng of total RNA from each sample was used to generate amplified and biotinylated complementary RNA (cRNA) from poly (A) RNA in a total RNA sample according to the user manual. IVT Incubation time was 16 hour. GeneAtlas® Hybridization, Wash and Stain Kit was used for hybridizing 3' IVT Array Strips according to the user manual (P/N 08-0306). Human genome array strips (HG-U219) were hybridized for 16 hours in a 45oC incubator, washed and stained and finally imaging was done with the GeneAtlas Fluidics and Imaging Station.


title: Transcriptional changes in HK-2 cells treated with lysophosphatidylcholine (LPC)

city: Nagaizumi-cho

first: Kentaro

last: Yoshioka

state: Shizuoka

address: 465-6, Shimotogari

institute: Kyowa Kirin Co. Ltd.

summary: To confirm gene expression changes upon LPC stimulation of HK-2 cells, we performed microarray analysis.

overall_design: Gene expression changes in HK-2 cells were measured at 24H after treated with 30, 60 µM of LPC (16:0) or (18:0).

first: Reiko

last: Inagi

first: Masaomi

last: Nangaku


title: circRNA expression in CRC and matched non-tumor tissues

city: Ningbo

first: Jipeng

last: Li

address: 251 East Baizhang Road

institute: The Affiliated People’s Hospital of Ningbo University

summary: To determine the circRNA expression profile in CRC and matched non-tumor tissues, we used circRNA microArray analysis form Arraystar to examine the expression of circRNAs in CRC and matched non-tumor tissues.

overall_design: Tissues were taken from patients undergoing CRC surgery were distributed into two experimental groups: normal tissue group and tumor tissue group, then perform circRNAs chips by Arraystar Human circRNAs chip (Arraystar)

first: Jian

last: Huang


title: cDNA Microarray 1: Crown developing xylem vs. Base developing xylem

summary: Transcriptional analysis comparing the transcriptomes of crown and base developing xylems from adult pine trees. Molecular factors responsible of wood properties can be identified by this strategy.

overall_design: Two-condition experiment Crown Differentiating Xylem vs. Base Differentiating Xylem. Biological replicates: 3 crown xylem, 3 base xylem, harvested from three different individual pine trees. Two technical replicates per array (A and Z). Two pairs of crown and base xylem RNA samples were labeled in the same way (crown xylem with Cy3 and base xylem with Cy5), while for labelling of the third pair dye-swap was performed (crown xylem labelled with Cy5 and base xylem with Cy3)


title: Establishment and characterization of ARK5 knockout gene in human myeloma  cell line generated by using CRISPR/Cas9 technology

status: Public on Oct 30 2021

city: Nagakute

first: Sivasundaram

last: Karnan

address: 1-1 Yazako karimata, Buid:2,　Room:304

department: biochemistry

institute: aichi medical university

summary: To investigate the effect of ARK5 on gene expression, we have employed whole genomic microarray expression profiling using KMS-11 and Sachi cells

overall_design: The KMS-11 and Sachi cells were plated at 80% confluence and cultured for 24 hours in vitro.

first: Akinobu

last: Ota


title: Initiation of organ maturation and fruit ripening in grapevine is controlled by the CARPO-NAC transcription factor

city: Paterna

first: José Tomás

last: Matus

state: Valencia

address: Carrer del Catedràtic Agustín Escardino Benlloch

institute: Institute for Integrative Systems Biology

summary: Grapevine is a woody temperate perennial plant and one of the most important fruit crops with global relevance in both the fresh fruit and winemaking industries. Unfortunately, global warming is affecting viticulture by altering developmental transitions and fruit maturation processes. In this context, uncovering the molecular mechanisms controlling the onset and progression of ripening could prove essential to maintain high-quality grapes and wines. Through a deep inspection of previously published transcriptomic data we identified the NAC family member VviCARPO (Controlled Adjustment of Ripening and maturation of Plant Organs) as a key regulator of grapevine maturation whose induction precedes the expression of well-known ripening associated genes. We explored VviCARPO binding landscapes through DAP-seq and overlapped its bound genes with transcriptomics datasets from stable and transient VviCARPO overexpressing grapevine plants to define a set of high-confidence targets. Among these, we identified key molecular ripening markers. Physiological, metabolic and promoter activation analyses showed that VviCARPO induces chlorophyll degradation and anthocyanin accumulation through the up-regulation of VviSGR1 and VviMYBA1, respectively, with the latter being up-regulated through a VviCARPO-VviNAC03 regulatory complex. Despite showing a closer phylogenetic relationship to senescent-related AtNAP homologues, VviCARPO complemented the nor mutant phenotype in tomato, suggesting it may have acquired a dual role as an orchestrator of both ripening- and senescence-related processes. Our data supports CARPO as a master regulator of the grapevine vegetative-to-mature phase organ transition and therefore an essential target for insuring fruit quality and environmental resilience.

overall_design:
Identification of binding sites for 3 transcription factors in Vitis vinifera by direct sequencing of affinity purified genomic DNA fragments.
Please note that the *_r1_r2.*events.narrowPeak file was generated from both r1, r2 replicates and is linked to the corresponding r1 sample records.

first: José

middle: T

first: Sara

last: Zenoni

first: Chiara

last: Foresti

first: Luis

last: Orduña


title: Gene expression profiling of HCT116 cells with or without PHGDH knockout

city: Shanghai

first: Yajuan

last: Zhang

address: 320 Yueyang Road

institute: Shanghai Institute of Biochemistry and Cell Biology

summary: HCT116 cells (WT or PHGDH KO) were harvested and mRNA were extracted for high-throughput sequencing

overall_design: WT v.s. PHGDH KO, in duplicate

first: Xiongjun

last: Wang


title: Transcriptional profiles LPC-HRas cells with or without  ETV4 knockdown

first: Xiaoli

last: Xu

address: Shanghai Jiaotong University

institute: Shanghai Jiaotong University

summary: To determine the molecular signaling pathways responsible for ETV4 regulation of cell growth, RNA sequencing was conducted on LPC-HRas cells with or without ETV4 knockdown

overall_design: Mouse liver tumor cells LPC-HRas were  infected with  the lentiviruses containing control shRFP or shETV4. The gene expression profiling in LPC-HRas cells with or without ETV4 knockdown was analyzed by RNA-seq.

first: Yongzhong

last: Liu


title: Pluripotent reprogramming arises from a unique subset of pre-disposed human dermal fibroblasts

city: Hamilton

first: Mickie

last: Bhatia

state: Ontario

address: 1200 Main Street West

department: Stem Cell and Cancer Research Institute (SCC-RI)

country: Canada

institute: McMaster University

summary: Induced cellular reprogramming to the pluripotent state offers a novel stem cell source for autologous transplantation. While recent studies have explored the role of factors required for induced pluripotent stem cell (iPSC) induction, the cellular and molecular basis of reprogramming from human fibroblasts remains elusive. Here, we have identified a subset of human dermal-derived fibroblasts that shares hallmark molecular and epigenetic features with pluripotent cells. Functional studies demonstrate that these cells contribute to the majority of human iPSCs generated from dermal fibroblasts and are dependent on heterogeneous fibroblast microenvironment for reprogramming competency. Molecular characterization indicated these predisposed fibroblasts were unique to other dermal derived stem cells and possessed features of proliferative selfrenewal. Our study reveals human fibroblasts are not equivalently capable of cellular reprogramming, and suggests that reprogramming factors overcome commitment steps that allow predetermined dermal fibroblasts to establish stable pluripotent state.

overall_design: Samples were obtained from human dermal fibroblasts (hFibs) transduced with a EOS lentiviral vector containing trimerized Oct4 enhancer elements that allow expression of GFP. Upon EOS transduction, we observed a small frequency of adult breast-derived dermal fibroblasts expressing GFP, fibroblasts were sorted based on GFP expression on FACS Aria (BD Pharmagin). 2 samples of GFP-positive and 2 samples of GFP-negative cells from transduced dermal fibroblasts and 1 sample of skin-derived precursors were obtained for this study.

first: Shravanti

last: Rampalli

first: Angelique

last: Schnerch

first: Eva

last: Szabo

first: Dong

middle: C

last: Wu

first: Marilyne

last: Levadoux-Martin

first: Aline

last: Fiebig

first: Olena

last: Morozova

first: Kausalia

last: Vijayaragavan

first: James

last: Ellis

first: Marco

last: Marra

first: Freda

last: Miller


title: RNA-Seq to validate the effect of substrate and format as drivers of transcriptional variance in human intestinal organoids

city: Houston

first: Noah

last: Shroyer

address: One Baylor Plaza, Room N710

department: Medicine Section of Gastroenterology and Hepatology

institute: Baylor College of Medicine

summary: An initial comparison of RNA-sequencing datasets from 251 experiments on 35 human intestinal organoid lines revealed that differences in transcriptional signatures could be influenced by variations in the format (3D, monolayers on transwells, and monolayers on 96-well plates) as well as substrate (collagen and Matrigel)  in which the organoid lines are cultured. While the organoid lines were a common to these datasets, there were differences in experimental approaches in different laboratories. To rule out potential confounding variables, we performed an RNA-Seq experiment using 5 jejunal organoid lines cultured at the same time in different formats and substrates to evaluate their roles in baseline transcriptional signatures and determine if key findings from the initial analysis could be validated.

overall_design: 25 samples were analyzed (no technical replicates). The cell lines used were J2,J3, J4, J6, J11. Each of these cell lines were plated as follows: 3D on Matrigel; Transwells on Matrigel; Transwells on collagen; Monolayers on Matrigel; Monolayers on collagen

first: Zachary

last: Criss

first: Sasirekha

last: Ramani

first: Mark

last: Estes

middle: F


title: Effect of substrate stiffness on human intestinal enteroids’ infectivity by enteroaggregative Escherichia coli

summary: Monolayer cultures of enteroids were cultured atop crosslinked poly(ethylene glycol)-based hydrogels that varied in stiffness as described in Manuscript. After the cells were adhered to the hydrogels, there were grown in differentiation medium for 5 days with medium changes every 48 h.

overall_design: Control samples from manuscript mentioned above. A total of 35 samples were analyzed of substrates of various stiffness

first: Ganesh

last: Swaminathan

first: Jane

last: Grande-Allen


title: Linc1548 promotes the transition of epiblast stem cells into neural progenitors by engaging OCT6 and SOX2

city: shanghai

first: Mingliang

last: Bai

state: shanghai

address: siping road

institute: tongji university

summary: The transition of embryonic stem cells from the epiblast stem cells (EpiSCs) to neural progenitor cells (NPCs), name as the neural induction process, is crucial for cell fate determination of neural differentiation. However, the mechanism of this transition is unclear. Here, we identified a long non-coding RNA (linc1548) as a critical regulator of neural differentiation of mouse embryonic stem cells (mESCs). Knockout of linc1548 did not affect the conversion of mESCs to EpiSCs, but delayed the transition from EpiSCs to NPCs. Moreover, linc1548 interacts with the transcription factors OCT6 and SOX2 forming an RNA-protein complex to regulate the transition from EpiSCs to NPCs. Finally, we showed that Zfp521 is an important target gene of this RNA-protein complex regulating neural differentiation. Our findings prove how the intrinsic transcription complex mediated by a lncRNA linc1548 and can better understand the intrinsic mechanism of neural fate determination.

overall_design: mRNA profiles of wild type (WT) and KO-Linc1548 46C cells at day 5 of neural differentiation were generated by microarray.


title: Genome-wide DNA methylation profiling of forensically relevant body fluids

status: Public on Oct 29 2021

city: Kashiwa

first: Ken

last: Watanabe

address: 6-3-1, Kashiwanoha

institute: National Research Institute of Police Science

type: Methylation profiling by genome tiling array

summary: Genome-wide DNA methylation profiling was performed to identify novel markers for DNA methylation-based identification of forensically forensically relevant body flulids. CpGs specifically methylated or unmethylated in saliva, vaginal swabs, blood and semen were searched by comparing beta values of about 850000 CpGs of pooled samples of each body fluid.

overall_design: Two sets of pooled samples of extracted DNA from each body fluid type (saliva, blood, semen, or vaginal swab) were prepared for genome-wide methylation analysis by Illumina Infinium MethylationEPIC. Each pooled DNA sample consisted of equal amounts of DNA from three individuals. Five hundred nanograms of each pooled sample were bisulfite-converted and analyzed according to the manual protocol of Illumina Infinium HD Methylation assay.


title: Telomere dysfunction instigates inflammation in inflammatory bowel disease

summary: Abstract: Inflammatory bowel disease (IBD) is a chronic inflammatory condition driven by diverse genetic and nongenetic programs that converge to disrupt immune homeostasis in the intestine. We have reported that, in murine intestinal epithelium with telomere dysfunction, DNA damage-induced activation of ataxia-telangiectasia mutated (ATM) results in ATM-mediated phosphorylation and activation of the YAP1 transcriptional coactivator, which in turn up-regulates pro-IL-18, a pivotal immune regulator in IBD pathogenesis. Moreover, individuals with germline defects in telomere maintenance genes experience increased occurrence of intestinal inflammation and show activation of the ATM/YAP1/pro-IL-18 pathway in the intestinal epithelium. Here, we sought to determine the relevance of the ATM/YAP1/pro-IL-18 pathway as a potential driver of IBD, particularly older-onset IBD. Analysis of intestinal biopsy specimens and organoids from older-onset IBD patients documented the presence of telomere dysfunction and activation of the ATM/YAP1/ precursor of interleukin 18 (pro-IL-18) pathway in the intestinal epithelium. Employing intestinal organoids from healthy individuals, we demonstrated that experimental induction of telomere dysfunction activates this inflammatory pathway.

overall_design: The transcriptome of undifferentiated Colonoids from 6 healthy and 6 Crohn's patients were analyzed for this study

first: Deepavali

last: Chakravarti

first: Ronald

last: DePinho


title: Enteroaggregative E. coli Adherence to Human Heparan Sulfate Proteoglycans Drives Segment and Host Specific Responses to Infection

summary: Enteroaggregative Escherichia coli (EAEC) is a significant cause of acute and chronic diarrhea, foodborne outbreaks, infections of the immunocompromised, and growth stunting in children in developing nations. There is no vaccine and resistance to antibiotics is rising. Unlike related E. coli pathotypes that are often associated with acute bouts of infection, EAEC is associated with persistent diarrhea and subclinical long-term colonization. Several secreted virulence factors have been associated with EAEC pathogenesis and linked to disease in humans, less certain are the molecular drivers of adherence to the intestinal mucosa. We previously established human intestinal enteroids (HIEs) as a model system to study host-EAEC interactions and aggregative adherence fimbriae A (AafA) as a major driver of EAEC adherence to HIEs. Here, we report a large-scale assessment of the host response to EAEC adherence from all four segments of the intestine across at least three donor lines for five E. coli pathotypes.

overall_design: Enteroids/colonoids on 96 well plate were generated as previously published (Poole, Rajan, & Maresso, 2018). The undifferentiated enteroids/colonoids monolayers were grown in CMGF(+) for 3 days and in differentiation media from days 3-5. The differentiated monolayers were then infected with bacterial cultures at a multiplicity of infection of 10 for 3 hours.

first: Anubama

last: Rajan

first: Anthony

last: Maresso


title: Transcriptomes of in vitro young leaves in eleven potato landraces

city: Ste-Anne-de-Bellevue

first: Martina

last: Stromvik

state: Quebec

address: 21,111 Lakeshore Rd

department: Department of Plant Science

institute: McGill University

summary: There is a need for expansion of the available potato genomic and transcriptomic resources in order to explore novel traits for potato improvement. Transcriptomic data derived from leaves from eleven native South American potato landraces (ten Peruvian and another; TBR Chilean) has been collected in order to aid the annotation of these genomes.

sample_organism:
Solanum tuberosum
Solanum tuberosum subsp. andigenum
Solanum x juzepczukii
Solanum phureja
Solanum chaucha
Solanum ahanhuiri
Solanum x curtilobum
Solanum stenotomum subsp. goniocalyx
Solanum stenotomum subsp. stenotomum

first: Noelle

last: Anglin

first: David

first: Maria

last: Kyriakidou

middle: V

last: Strömvik

first: Saireddy

last: Achakkagari

first: Ilayda

last: Bozan

first: Helen

last: Tai


title: Spatial alanine metabolism determines local growth dynamics of Escherichia coli colonies

city: Marburg

first: Francisco

last: Diaz-Pascual

address: Karl-von-Frisch-Straße 10

institute: Max Planck Institute for Terrestrial Microbiology

summary: By using transcriptomics, we studied the spatiotemporal transcriptional changes in Escherichia coli biofilm colonies

overall_design: Whole colony measurenments were performed on colonies grown for 12 h to 72 h in triplicate (samples 1-24). For colonies grown for 72 h, the transcriptional profile of anoxic and oxic cells was analyzed using FACS and RNAseq with 4 replicates (samples  41-48). Samples 25-40 are controls

last: DiazPascual

first: Kazuki

last: Nosho

first: Knut

last: Drescher


title: Impaired Skeletal Muscle Repair in Healthy Young Adults with Type 1 Diabetes Mellitus

first: Athan

middle: G

last: Dial

state: ON

address: 1280 Main Street West

department: Medical Sciences

summary: Preclinical models of type 1 diabetes mellitus exhibit marked declines in skeletal muscle health including significant impairments in muscle repair. The present study investigated, for the first time, whether muscle repair was altered in young adults with uncomplicated type 1 diabetes (T1D) following damaging exercise.In this cohort study, eighteen physically-active young adults (M=22.1, SEM=0.9 years) with T1D (n, male/female=4/5; MHbA1c= 58, SEMHbA1c=5.9 mmol/mol) and without T1D (n, male/female=4/5) performed 300 unilateral eccentric contractions (90°s-1) of the knee extensors. Prior to exercise, at 48-hours and at 96-hours after exercise, participants gave a venous blood sample and vastus lateralis biopsy and performed a maximal voluntary isometric knee extension. Skeletal muscle extracellular matrix content, and satellite cell content/proliferation were assessed by immunofluorescence. Transmission electron microscopy was used to quantify ultrastructural damage.Maximal isometric strength was comparable between T1D and their sex-matched control group prior to exercise for both sexes. Immediately following damaging exercise, strength was decreased in both groups but there was a moderate effect size for lower strength during recovery in the T1D group at both 48-hours and 96-hours. Serum creatine kinase, an indicator of muscle damage, was moderately higher in T1D participants compared to controls at rest, and exhibited a small elevation 96-hours after exercise. Immunofluorescence analyses showed satellite cell content was lower at all timepoints in those with type 1 diabetes. T1D participants demonstrated a moderate delay in satellite cell proliferation (as assessed by Pax7+/Ki67+ nuclei counts) after exercise, reaching peak levels of proliferation 48-hours after control participants. Despite these differences, those with T1D did not exhibit greater ultrastructural muscle damage than controls as assessed by electron microscopy. Finally, a transcriptomic investigation of T1D muscle revealed several networks of dysregulated genes involving RNA translation as well as mitochondrial respiration function, providing potential explanations for previous observations of mitochondrial dysfunction in similar T1D cohorts. Our novel findings indicate that skeletal muscle from physically active, young adults with moderately controlled T1D may have a reduced ability to handle, and repair from, damaging exercise. While larger cohort studies are clearly needed, these results suggest that those with T1D may require longer recovery times following damaging exercise.

overall_design: 16 samples were analysed. We generated the following pairwise comparisons using Transcriptome Analysis Console: T1D>CON (PRE); T1D>CON (48H); T1D>CON (Interaction, PRE to 48H). Gene set enrichment analysis was performed on each comparison. Gene sets with FDR ≤ 10% were included in final analyses.

first: Thomas

middle: J

last: Hawke


title: The antifolate pemetrexed induces innate tolerance in human macrophages

status: Public on Oct 28 2021

city: Madrid

first: Amaya

middle: Rocío

last: Puig Kröger

state: Madrid

address: Dr. Esquerdo 46

department: Immunology

institute: Hospital General Universitario Gregorio Marañón

summary: Macrophage phenotypic and functional heterogeneity derives from tissue-specific transcriptional signatures shaped by the local microenvironment. GM-CSF drives the generation of human monocyte-derived macrophages with a potent pro-inflammatory activity upon stimulation. One-carbon metabolism (OCM) is a complex network of biosynthetic pathways that includes de novo biosynthesis of purines and thymidylate, amino acid metabolism, and methylation reactions. We explored the molecular impact of blocking OCM with the anti-folate pemetrexed (PMX) on the gene expression profile in LPS-activated GM-CSF-primed human monocyte derived macrophages. The specificity of blocking OCM with the anti-folate PMX was demonstrated with folinic acid.

overall_design: mRNA profiles of human monocyte-derived macrophages exposed to 10 ng/ml LPS for 3h. Monocytes were cultured for 7 days containing GM-CSF (1000 U/ml) to generate GM-CSF-polarized macrophages (GM-MØ). GM-CSF was added every two days. Pemetrexed (PMX, 50 nM), folinic acid (FA, 5 µM) or both (PMX+FA) were added once on monocytes together with GM-CSF.

first: Sara

last: Romero

first: Raquel

last: Campos

middle: P

last: Kröger


title: Application of the 3’ mRNA-Seq using unique molecular identifiers in highly degraded RNA derived from formalin-fixed, paraffin-embedded tissue

city: Rochester

first: Jin Sung

last: Jang

address: 200 1st st sw

institute: Mayo Clinic

summary:
Background: Archival formalin-fixed, paraffin-embedded (FFPE) tissue samples with clinical and histological data are a singularly valuable resource for developing new molecular biomarkers. However, transcriptome analysis remains challenging with standard mRNA-seq methods as FFPE derived-RNA samples are often highly modified and fragmented. The recently developed 3’ mRNA-seq method sequences the 3’ region of mRNA using unique molecular identifiers (UMI), thus generating gene expression data free from PCR amplification bias. In this study, we evaluated the performance of the 3’ mRNA-Seq using Lexogene QuantSeq 3’ mRNA-Seq Library Prep Kit FWD with UMI, comparing with TruSeq stranded mRNA-Seq and RNA Exome Capture. The fresh-frozen (FF) and FFPE tissues yielded nucleotide sizes range from 13% to >70% of DV200 values; input amounts ranged from 1ng to 100ng for validation.
Results: The total mapped reads of the 3’ mRNA-Seq to the reference genome ranged from 99% to74% across all the samples. After PCR bias correction, total mapped reads ranged from 3% to 56%. The 3’ mRNA-Seq data showed highly reproducible data in the UHR (R>0.94) at different input amounts from 1ng to 100ng, and FF and FFPE paired samples (R=0.93) at 10 ng.  Severely degraded FFPE RNA with 13% to 30% of DV200 value showed good concordance (R>0.90) with 100ng input. A moderate correlation was observed when directly comparing  3’ mRNA-Seq data with TruSeq stranded mRNA-Seq (R=0.78) and RNA Exome Capture data (R>0.67).
Conclusion: In this study, 3’ mRNA-Seq with PCR bias correction using UMI is a suitable method for gene quantification in both FF and FFPE RNAs. The 3’ mRNA-Seq may be applied to severely degraded RNA from FFPE tissues and generate high-quality sequencing data.

overall_design: The 3’ mRNA-Seq using Lexogene QuantSeq 3’ mRNA-Seq Library Prep Kit FWD with UMI is compared with TruSeq stranded mRNA-Seq and RNA Exome Capture kits. The fresh-frozen (FF) and FFPE tissues yielded nucleotide sizes range from 13% to >70% of DV200 values; input amounts ranged from 1ng to 100ng for validation.

first: Jin

middle: S


title: Human monocyte-derived macrophages polarized by GM-CSF in the presence or absence of 50 nanomolar Pemetrexed

summary: Macrophage phenotypic and functional heterogeneity derives from tissue-specific transcriptional signatures shaped by the local microenvironment. GM-CSF drives the generation of human monocyte-derived macrophages with a potent pro-inflammatory activity upon stimulation. One-carbon metabolism (OCM) is a complex network of biosynthetic pathways that includes de novo biosynthesis of purines and thymidylate, amino acid metabolism, and methylation reactions. We explored the molecular impact of blocking OCM with the anti-folate pemetrexed (PMX) on the gene expression profile in GM-CSF-primed human monocyte derived macrophages.

overall_design: mRNA profiles of human macrophages differentiated with GM-CSF (GM-MØ) from peripheral blood monocytes in the absence or in the presence of the antifolate pemetrexed (PMX).


title: A network-based approach reveals the dysregulated transcriptional regulation in non-alcohol fatty liver disease

status: Public on Oct 27 2021

city: Solna

first: Hong

last: Yang

state: Stockholm

address: Tomtebodavägen 23A

department: Systems Biology

country: Sweden

institute: KTH Royal Institute of Technology/Science For Life Laboratory

summary: Non-alcohol-related fatty liver disease (NAFLD) is a leading cause of chronic liver disease worldwide. We performed network analysis to investigate the dysregulated biological processes in the disease progression and revealed the molecular mechanism underlying NAFLD. Based on network analysis, we identified a highly conserved disease-associated gene module across three different NAFLD cohorts and highlighted the predominant role of key transcriptional regulators associated with lipid and cholesterol metabolism. In addition, we revealed the detailed metabolic differences between heterogenous NAFLD patients through integrative systems analysis of transcriptomic data and liver-specific genome-scale metabolic model. Furthermore, we identified transcription factors (TFs), including SREBF2, HNF4A, SREBF1, YY1 and KLF13, showing regulation of hepatic expression of genes in the NAFLD-associated modules and validated the TFs using data generated from a mouse NAFLD model. In conclusion, our integrative analysis facilitated our understanding of the regulatory mechanism of these perturbed TFs and associated biological processes.

overall_design: C57BL/6J mice were fed a standard mouse chow diet and housed in a 12‐h light–dark cycle. From the age of 8 weeks, the mice were then divided into two groups of 5 mice fed with chow diet, 4 mice fed with high-sucrose diet for 3 weeks, respectively. At the age of 11 weeks, we performed a genome-wide transcriptomic analysis on tissue samples obtained from these mice.

first: Muhammad

last: Arif

first: Meng

last: Yuan

first: Xiangyu

last: Li

first: Ko

middle: E

last: Shong

first: Hasan

last: Turkez

first: Jens

last: Nielsen

first: Mathias

last: Uhlén

first: Jan

last: Borén

first: Cheng

last: Zhang

first: Adil

last: Mardinoglu


title: Immunomagnetic B cell isolation as a tool to study blood cell subsets and enrich B cell transcripts

city: Bethesda

first: Amanda

last: Henning

state: MD

address: 9000 Rockville Pike

institute: National Institutes of Health

summary:
Objective: Transcriptional profiling of immune cells is an indispensable tool in biomedical research; however, heterogenous sample types routinely used in transcriptomic studies may mask important cell type-specific transcriptional differences. Techniques to isolate desired cell types are used to overcome this limitation. We sought to evaluate the use of immunomagnetic B cell isolation on RNA quality and transcriptional output. Additionally, we aimed to develop a B cell gene signature representative of a freshly isolated B cell population to be used as a tool to verify isolation efficacy and to provide a transcriptional standard for evaluating maintenance or deviation from traditional B cell identity.
Results: We found RNA quality and RNA-sequencing output to be comparable between donor-matched PBMC, whole blood, and B cells following negative selection by immunomagnetic B cell isolation. Transcriptional analysis enabled the development of an 85 gene B cell signature. This signature effectively clustered isolated B cells from heterogeneous sample types in our study and naïve and memory B cells when applied to transcriptional data from a published source. Additionally, by identifying B cell signature genes whose functional role in B cells is currently unknown, our gene signature has uncovered areas for future investigation.

overall_design: mRNA profiling of freshly collected whole blood, peripheral blood mononuclear cells (PBMC), and isolated B cells from n=4 healthy donors (human).

first: Valeria

last: De Giorgi


title: Gene structure, differential exon usage, and expression of the testis long intergenic non-protein coding RNA 1016 in humans reveals isoform-specific roles in controlling biological processes

city: El Paso

first: Shrikanth

last: Gadad

state: TX

address: 5001 El Paso Drive

department: Department of Molecular and Translational Medicine

institute: Texas Tech University Health Sciences Center El Paso

summary: Long noncoding RNAs (lncRNAs) have emerged as critical regulators of biological processes. The constant expansion of newly-identified lncRNA genes requires that each one be comprehensively annotated to understand its molecular functions. Here, we describe a detailed characterization of the gene which encodes long intergenic non-protein coding RNA 01016 (LINC01016, a.k.a., LncRNA1195) with a focus on its structure, exon usage, and expression in human and macaque tissues. In this study, we show that it is exclusively conserved among non-human primates, suggesting its recent evolution and is expressed and processed into 12 distinct RNAs in testis, cervix, and uterus tissues. Further, we integrate de novo annotation of expressed LINC01016 transcripts and isoform-dependent gene expression analyses to show that human LINC01016 is a multi-exon gene, processed through differential exon usage with isoform-specific functions. Furthermore, in gynecological cancers, such as cervical squamous cell carcinoma and uterine corpus endometrial carcinoma, LINC01016 is downregulated; however, its higher expression is predictive of relapse-free survival in these cancers. Collectively, these analyses reveal that, unlike coding RNAs, lncRNA genes are tightly regulated and precisely processed to generate isoforms with distinct biological roles in specific tissues.

overall_design: Six different transcripts of the LINC01016 human gene were cloned in a mammalian expression vector and sequenced. Each transcript was expressed in a heterologous cell line (HEK293 cells, LINC01016 is not transcribed).

first: Enrique

middle: I

last: Ramos

first: Barbara

first: Yasmin

last: Vasquez

first: Ken

middle: Y

last: Lin

first: Ramesh

last: Choudhari


title: Transcriptomic study of E. coli 042 with a variant of aggR gene with a FRT sequence inserted in its 3´UTR by RNA-seq

city: Barcelona

first: Mario

last: Huttener Queiroz

address: Av. Diagonal 643

institute: Universitat de Barcelona

summary: A common genomic feature of most EAEC strains is the presence of a virulence plasmid termed pAA. Plasmid-encoded virulence determinants are, among others, a transcriptional activator termed AggR, a member of the AraC-XylS family of transcription factors. We have previously determined the direct correlation between (p)ppGpp, expression of AggR and biofilm development in strain EAEC 042 (https://doi.org/10.3389/fmicb.2018.00717). In this work we characterize a novel variant of the aggR gene. We modified its 3´UTR by insertion of a FRT sequence, which have generated a series of different phenotypes. We used RNA-seq to compare the transcriptome of the wt strain and its aggR3UTRFRT variant grown at 37ºC in LB medium.

overall_design: E. coli 042 and derivative aggR3UTRFRT mRNA profiles were generated by Illumina RNA-seq. RNA extraction, DNase treatment, evaluation of RNA quality and cDNA libraries for Illumina sequencing were performed by Vertis Biotechnologie AG, Freising-Weihenstephan, Germany. Bacterial cells were grown until O.D.600 of 2.0 in LB medium at 37ºC with 200rpm of agitation. Three replicates were grown from each strain, 1 ml of each replicate was collected and bacterial pellets were maintained at -80ºC. Bacterial pellets were sent to Vertis Biotechnologie AG, Freising-Weihenstephan, Germany in dry ice. E. coli 042 wt was used as genome reference (NC_017626 for the chromosome and NC_017627 for the pAA plasmid) for analysis.

first: A

last: Prieto

first: M

last: Hüttener

last: Juárez


title: Pharmacogenomics characterization of the MDM2 inhibitor MI-773 reveals candidate tumours and predictive biomarkers

city: Freiburg

first: Vincent

last: Vuaroqueaux

address: Am Flughafen 14

department: Bioinformatics

institute: 4HF Biotec

summary: Affymetrix SNP6.0  array data used to reveal predictive biomarkers of tumour sensitivity to MI-773 . MI-773 is a recently developed small-molecule inhibitor of the Mouse Double Minute 2 (MDM2) proto-oncogene. Here we report an integrative pharmacogenomic study to gain further insights into the therapeutic potential of the compound

overall_design:
Affymetrix SNP6.0  arrays were performed according to the manufacturer's directions on DNA extracted from cryopreserved materials.
Copy number analysis of Affymetrix SNP6 arrays was performed for 139 cell lines.

first: Hans

middle: R

last: Hendriks

first: Hoor

last: Al-Hasani

first: Anne-Lise

last: Peille

first: Samayita

last: Das

first: Heinz-Herbert

middle: H

last: Fiebig


title: Activation of proneuronal transcription factor Ascl1 in maternal liver ensures a healthy pregnancy

city: Indianapolis

first: Guoli

last: Dai

state: IN

address: 723 W Michigan St. SL306

department: Biology

institute: Indiana University Purdue University Indianapolis

summary:
Background & Aims: Maternal liver exhibits robust adaptations to pregnancy to accommodate the metabolic needs of developing and growing placenta and fetus by largely unknown mechanisms. We found that achaete-scute homolog-like 1 (Ascl1), a basic helix-loop-helix transcription factor essential for neuronal development, is highly activated in maternal hepatocytes during the second half of gestation in mice.
Methods: To investigate whether and how Ascl1 plays a pregnancy-dependent role, we deleted the Ascl1 gene specifically in maternal hepatocytes from mid-gestation until term.
Results: As a result, we identified multiple Ascl1-dependent phenotypes. Maternal livers lacking Ascl1 exhibited aberrant hepatocyte structure, increased hepatocyte proliferation, enlarged hepatocyte size, reduced albumin production, and elevated release of liver enzymes, indicating maternal liver dysfunction. Simultaneously, maternal pancreas and spleen and the placenta displayed marked overgrowth; and the maternal ceca microbiome showed alterations in relative abundance of several bacterial subpopulations. Moreover, litters born from maternal hepatic Ascl1-deficient dams experienced abnormal postnatal growth after weaning, implying an adverse pregnancy outcome. Mechanistically, we found that maternal hepatocytes deficient for Ascl1 exhibited robust activation of insulin-like growth factor 2 expression, which may contribute to the Ascl1-dependent phenotypes widespread in maternal and uteroplacental compartments.
Conclusion: In summary, we demonstrate that maternal liver, via activating Ascl1 expression, modulates the adaptations of maternal organs and the growth of the placenta to maintain a healthy pregnancy. Our studies reveal Ascl1 as a novel and critical regulator of the physiology of pregnancy.

overall_design: Total RNA was isolated from livers of WT control (Ascl1fl/fl; n=4) and hepatocyte-specific Ascl1 KO (hep-Ascl1-/-; n=5) mice on gestation day 15 and was subjected to RNA-sequencing

first: Joonyong

last: Lee

first: Veronica

last: Garcia


title: Pharmacogenomic characterization of the MDM2 inhibitor MI-773 reveals candidate tumours and predictive biomarkers

summary: MI-773 is a recently developed small-molecule inhibitor of the Mouse Double Minute 2 (MDM2) proto-oncogene. Here we report an integrative pharmacogenomic study to gain further insights into the therapeutic potential of the compound.

overall_design: Total RNA samples were extracted using the mirVana kit (Ambion) following the manufacturer's recommendations. After standard quality controls RNA were subjected to Affymetrix HGU133 Plus2.0 microarray hybridization.